Despite considerable effort and progress in developing tetravalent vaccines against the four serotypes of dengue virus (DENV), one of the major unmet challenges is the difficulty in eliciting balanced neutralizing (NT) antibodies (Abs) against all four serotypes and to lower the risk of antibody-dependent enhancement (ADE), mediated mainly by cross-reactive and weakly or non-NT Abs. Studies of human Abs after DENV infection have shown the immunodominance of cross-reactive and weakly or non-NT Abs recognizing the fusion loop (FL) of envelope (E) protein over the type-specific and potent NT Abs, and the presence of cross-reactive and weakly or non-NT anti-precursor membrane (prM) Abs. Whether such immunodominance can be modulated to induce potent NT Abs without cross-reactive and weakly or non-NT Abs remains unknown. We recently found a DNA vaccine expressing mature DENV particles induced potent NT Abs with minimal cross-reactive, weakly or non-NT Abs, suggesting that specific modulation of DENV particles might induce superior Ab responses. Our long-term goal is to develop a safe and effective DENV vaccine. The objective of the proposed research is to understand the immunodominance of DENV E protein and to test whether mature or FL-modified mature DENV particles can induce potent NT Abs without cross-reactive, weakly or non-NT Abs. The central hypothesis is that mature DENV particles induce potent NT Abs, less anti-FL Abs and no anti-prM Abs, thus reducing the risk of ADE compared with mixed DENV particles. The objective will be achieved by the three specific aims: The first specific aim is to define the E protein epitopes and the accessibility, binding avidity and NT potency of a large panel of anti-E mAbs on mature, mixed and immature DENV particles. We hypothesize that differential epitope accessibility and binding avidity of anti-E Abs to different DENV particles may account for the immunodominance of E protein. The second aim is to demonstrate the superiority of DNA vaccines expressing mature DENV particles, over DNA vaccines expressing mixed particles, in eliciting potent NT Abs and minimal potential enhancing Abs and providing protection in outbred mice and AG129 mice, a well-established dengue murine model. The third aim is to demonstrate the superiority of DNA vaccines expressing FL-modified mature DENV particles, over DNA vaccines expressing non-modified mature DENV particles, in eliciting potent NT Abs and minimal potential enhancing Abs and providing protection in murine models. The significance of the proposed research rests on its detailed understanding of the immunodominance of DENV E protein and on the demonstration that such immunodominance can be modulated by mature and FL-modified DENV particles to induce potent NT Abs and minimal infection-enhancing Abs. This represents a novel strategy for DENV vaccine design and cannot be achieved by all current DENV particle-based candidate vaccines. The proposed research can be translated to several more advanced DENV vaccine candidates as the second generation of safe and effective DENV vaccines.